疏绵状嗜热丝孢菌的糖基化热稳定锰超氧化物歧化酶及真菌锰超氧化物歧化酶的系统学（英文）  

A glycosylated thermostable manganese superoxide dismutase from Thermomyces lanuginosus and phylogeny of fungal manganese superoxide dismutases

作　　者：张佳[1] 李多川[1] ZHANG Jia;LI Duo-Chuan(Department of Mycology, Shandong Agricultural University, Tai＇an, Shandong 271018, China)

机构地区：[1]山东农业大学菌物学系,山东泰安271018

出　　处：《菌物学报》2018年第6期722-736,共15页Mycosystema

基　　金：Supported by the Chinese National Key Technology Support Program(2015BAD15B05);the National Natural Science Foundation of China(31571949);National High-Technology Research&Development Program of China(2012AA10180402)

摘　　要：疏绵状嗜热丝孢菌Thermomyces lanuginosus是一种广泛分布的嗜热真菌,产生的酶在高温条件下具有高活性和稳定性。从该嗜热真菌中克隆了一种锰超氧化物歧化酶(manganese superoxide dismutase,MnSOD)基因Tlmnsod1(EU364837),cDNA全长791bp,开放阅读框654bp,编码217个氨基酸,没有信号肽和前导肽序列。Tlmnsod1基因在酵母中高效表达,经纯化获得了电泳均一分子量约为27kDa的重组蛋白TlMnSOD1。表达的TlMnSOD1酶是糖基化蛋白,具高的热稳定性,最适反应pH和温度分别为8.0和55℃。通过对真菌的40种锰超氧化物歧化酶的细胞定位和系统学分析可知:真菌的锰超氧化物歧化酶可以分为胞内MnSOD、线粒体MnSOD和胞外MnSOD;锰超氧化物歧化酶可以作为分子标记来研究真菌的进化关系;3种嗜热真菌MnSOD分别与非嗜热真菌MnSOD聚类在3个不同的组中。本研究为MnSOD的糖基化和作为分子标记用于真菌种的鉴定提供了证据。A manganese superoxide dismutase gene（Tlmnsod1） from Thermomyces lanuginosus was cloned and expressed in Pichia pastoris. The full-length cDNA of the MnSOD was 791 bp and contained a 654 bp open reading frame encoding 217 amino acid residues. No signal peptide or leader peptide was present in the putative amino acid sequence of TlMnSOD1. The enzyme belongs to the Mn/Fe-binding family. The highly conserved motif of the MnSOD family, DXWEHAYY, was found in TlMnSOD1. The gene was transformed into Pichia pastoris. The recombinant TlMnSOD1 was shown to be a glycosylated MnSOD. The enzyme exhibited optimal activity at 55℃ and pH 8.0 and had high thermostability. Analysis of subcellular localization and phylogeny of the TlMnSOD1 and 39 other fungal MnSODs from Ascomycota predicted three kinds of MnSODs（mitochondrial, cytoplasmic, and extracellular MnSODs）. Each of three MnSODs from thermophilic fungi was clustered together with non-thermophilic mesophilic fungi in different groups in the phylogeny tree. The results of this study provide evidences that MnSOD may be glycosylated and used as a molecular marker for identification of closely related fungal species.

关 键 词：疏绵状嗜热丝孢菌 糖基化锰超氧化物歧化酶 热稳定性 系统学 分子标记 

分 类 号：Q936[生物学—微生物学]